Gallstone disease affects more than 30,000,000 Americans and results in more than 750,000 cholecystectomies in the United States annually. Gallstones can cause extrahepatic cholestasis by occluding the common bile duct. In bile, bile acids and phospholipids are important for the solubilization of cholesterol. When the composition of bile changes such that the ratio of bile acids to cholesterol decreases (either decreased bile acid synthesis and/or increased cholesterol secretion into bile), the probability for formation of cholesterol gallstones increases. The broad objective of this proposal is to understand how the transcription factor Nuclear Factor-E2-related factor 2 (Nrf2) regulates cholesterol and bile acid metabolism and disposition during cholestasis and during exposure to a high cholesterol diet. Preliminary studies show that liver transporter expression is different in livers from wild-type (WT) mice and Nrf2-null mice during cholestasis. Thus, Specific Aim 1 will expand upon these initial findings and determine whether drug disposition differs between WT and Nrf2-null mice during cholestasis. Importantly, preliminary studies also demonstrated that bile acid levels are decreased in liver and serum from Nrf2-null mice as compared to WT mice, suggesting that Nrf2 is important for bile acid synthesis. Therefore, Specific Aim 2 will determine the mechanism by which Nrf2 regulates bile acid levels in liver. Preliminary data also indicated that Nrf2-null mice exhibit increased formation of cholesterol gallstones when exposed to a high cholesterol diet, and studies in Specific Aim 3 will determine the mechanism by which Nrf2-null mice are more susceptible to gallstone formation. Studies in aim 3 will examine biliary cholesterol, bile acid, and phospholipid secretion as well as examine differences in heptic expression of genes for cholesterol and bile acid synthesis, metabolism, and disposition. Together, the proposed studies will provide novel information regarding liver regulation of cholesterol and bile acids and provide valuable insight into the pathogenesis of gallstone formation.